Human IgG Fc-engineering for enhanced plasma half-life, mucosal distribution and killing of cancer cells and bacteria

Monoclonal IgG antibodies constitute the fastest growing class of therapeutics. Thus, there is an intense interest to design more potent antibody formats, where long plasma half-life is a commercially competitive differentiator affecting dosing, frequency of administration and thereby potentially patient compliance. Here, we report on an Fc-engineered variant with three amino acid substitutions Q311R/M428E/N434W (REW), that enhances plasma half-life and mucosal distribution, as well as allows for needle-free delivery across respiratory epithelial barriers in human FcRn transgenic mice. In addition, the Fc-engineered variant improves on-target complement-mediated killing of cancer cells as well as both gram-positive and gram-negative bacteria. Hence, this versatile Fc technology should be broadly applicable in antibody design aiming for long-acting prophylactic or therapeutic interventions.

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Data exclusions
Publicly available data sets used: Protein database accession number 4NOU.
Protein database accession number 1HZH.
The ex vivo human placental perfusion study involves the use of human biological material.Sex was considered during study design and only pregnant female participants were enrolled since they are the only sex that have placentas.Sex was determined based on assignment.Gender was not considered as part of the study design.
As only females participated in the study disaggregated data for sex and gender was not collected.
Sex or gender was not considered during collection of human blood samples prior to isolation of human immune cells used for cellular assays.
Race, ethnicity or other social groupings were not considered as part of the study design.
To minimize variation in the ex vivo placenta perfusion model, mothers who smoked, had diabetes or other pregnancy complications were excluded from the study.
Placentas from uncomplicated pregnancies resulting in vaginal or caesarean section were donated by women givin birth at Copenhagen University Hospital.Informed consent was obtained in accordance with the Declaration of Helsinki.
Human blood samples were collected following informed consent at Utrecht Medical Center or University Medical Center Schleswig-Holstein.
No self-selection bias or other biases have been recorded. -

Antibodies
Antibodies used Validation using Grubb`s test.This occurred only in a limited subset of data points and had no impact on the conclusions.
Analysis of samples from in vivo studies were performed twice using established methodology and reviewed by internal and external experts.Cellular experiments were performed 2 or 3 times as independent experiments using 2-4 technical replicates.Biochemical experiments were repeated at least once and analyzed using appropriate evaluation software.All stated instrumentation underwent routinely maintenance and quality assessment both prior to and during the data collection period.All attempts at replication were successful.
Not relevant to the study, as it would compromise ability to interpret data.
Irrelevant to our study, as it would compromise the ability to interpret data.
-Anti-Human IgG (Fc-specific) produced in goat, Sigma, I2136, 0000154675.Not relevant to to the study.
Not relevant to to the study.
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Sample size in animal experiments were determined based on ethical perspectives (the 3 Rs) and previously published and collected data confirming the sufficiency of the used sample size for robust analysis and statistical analysis.Animal experiments: PMID: 35982144, PMDI: 35118359, PMID: 29391560.Human placenta perfusion experiment: PMID: 35981406, PMID: 23843496.Cellular experiments: PMID29391560, PMID: 26718855.Data was excluded from further interpretation if found to be below the sensitivity threshold of the analytical method or identified as outlayers nature portfolio | reporting summary